CHARGED LIPOSOME PREPARATION

Abstract: The invention relates to a method of administering a liposome preparation having an effective amount of an electrically charged component which is effective in reducing adverse effects of a liposome preparation, e.g., in reducing hemodynamic effects, and improving host tolerance.

Description: CHARGED LIPOSOME PREPARATION Background of the Invention This invention relates to liposomes and their administration to host organisms. Liposomes have been used as carriers for active agents, including therapeutic, diagnostic, and prophylactic agents. Liposomes generally comprise amphipathic compounds, having a hydrophilic head and a hydrophobic tail, which react under certain conditions to form a bimolecular leaf structure of at least two layers of lipid, in which the polar head groups are at the interface between the aqueous medium and the lipid and the hydrophobic tails interact to form an environment that excludes water. The lipid bilayers are stable structures held together by the non-covalent interaction of the hydrocarbon groups of the acyl groups. When the lipid bilayer closes in on itself, it forms a spherical vesicle called a liposome having an internal space separated from the external environment. Desired agents may be encapsulated in this space, or within the bilayer, itself. The lipids which comprise the liposome are typically phosphoglycerides, such as phosphatidyl choline (lecithin) , and sphingolipids. Liposomes may be used as carriers for, e.g., thera¬ peutic, diagnostic, and prophylactic agents. They are especially advantageous in that they may protect suscep¬ tible agents from degradation, thereby increasing the time during which the agent is active in the body. In addition, tissue or cell specific targeting of the lipo¬ some may be achieved by incorporating a component into the liposome, e.g., coupling, conjugating, absorbing or adsorbing it to the liposome bilayer surface, which is selective for a specific tissue or cell type. For example, lipids, antibodies, lectins, receptors, ligands, and other such components may be incorporated into liposomes for the purpose of achieving tissue specific targeting. A further application of liposome technology is in the treatment, prophylaxis, and diagnosis of liver disorders. In spite of the large variety of imaging techniques and modalities currently available, accurate assessment of liver lesions remains a difficult diagnostic problem. One of the reasons for the diagnostic difficulties is the low contrast difference between normal and tumor liver parenchyma, which only allows delineation of lesions above 1 to 2 cm. Accordingly, it would be desirable to be able to concentrate a radiographic or MRI contrast medium in the liver in order to increase the contrast for a time period that permits thorough examination of the entire organ. One way to achieve uptake of hydrophilic substances into the liver is to encapsulate them in liposomes. After i.v. administration, liposomes may be taken up by the phagocytic cells of the reticulo-endothelial system. Thus, they are chiefly concentrated in the Kupffer-cells of the liver and macrophages in the spleen. Since phagocytic activity is not displayed by tumor tissue, the liposomal contrast agent can be concentrated selectively in the healthy tissue. This results in an increased density difference (Δ HU for X-ray imaging, ΔT1 or ΔT2 for MRI) . Various investigators have been able to demonstrate the effectiveness of such an approach in principle. In most cases, however, they have been unable to achieve a sufficient density difference due to the low iodine- loading capacity of their liposomes; nor have any of the methods of preparation for the contrast agent carrying liposomes applied proven suitable for reproducible large- scale production of pharmaceutically acceptable liposome preparations (adequate shelf-life, sterile, pyrogen free, high iodine encapsulation) . A method of preparation of contrast-carrying lipo¬ somes has been developed which overcomes all the above limitations. See WO 91/16039; P 40 135802. This process called "ethanol-evaporation method" enables reproducible production of large amounts of liposomes under aseptic conditions. The original liposome suspension produced by this method is lyophilized to obtain a product with an adequate shelf life. Prior to application, the lyophili- zate is resuspended with 135 mM mannitol solution to give a liposomal suspension which is infused after a filtration step. Using contrast-carrying liposomes produced by this method, opacification of the liver and spleen has been achieved. On the basis of initial biodistribution studies, iopromide was chosen as liposomal contrast agent because its 7-day retention values in the liver and spleen are significantly lower than those of iotrolan. Although liposomes are potentially an important advance in drug delivery systems, such as for carrying contrast media as described above, their use in living organisms is limited owing to adverse systemic side effects which may be observed upon liposome administration. These systemic effects include, e.g., transient hemodynamic depression, affecting blood pressure, heart rate, and ECG; depression of blood counts. For example, it is known that liposomes may have adverse hemodynamic effects when administered intravenously to animals, posing serious hazards to the safety of the host animal. In view of the occurrence of such systemic effects, it may not be possible to use such liposome preparations for drug delivery. Surprisingly, it has been found that when these same liposomes are prepared but incorporating an electrically-charged component the adverse systemic effects upon administration are reduced or eliminated. Brief Description of the Drawing Figure 1 is a schematic diagram of the ethanol- evaporation method of making liposomes. Figure 2 is mean blood pressure (percent of prevalue) following intravenous infusion of liposomes made from different lipids (300 mg lipid/kg, 100 mg lipid/kg/min) and mannitol (300mM) , n=6, means+SEM. • Mannitol O DSPC/DSPG D DSPC Figure 3 is total peripheral resistance (percent of prevalue) following intravenous infusion of liposomes made from different lipids (300 mg lipid/kg, 100 mg lipid/kg/min) and mannitol (300mM) , n=6, means+SEM. • Mannitol O DSPC/DSPG D DSPC Figure 4 is enddiastolic pressure (EDP) following intravenous infusion of liposomes made from different lipids (300 mg lipid/kg, 100 mg lipid/kg/min) and mannitol (300mM) , n=6, means+SEM. • Mannitol o DSPC/DSPG D DSPC Summary of the Invention By incorporating an electrically charged component into liposomes which have an adverse systemic effect, it has been found that the resulting liposomes reduce or eliminate the occurrence of undesired or adverse systemic effects which are otherwise observed when the liposomes are formed from the same lipid constituents but omitting the electrically charged component are administered. In particular, the addition of a component having a negative charge, e.g., a fatty acid such as stearic acid, to a liposome preparation improves the host's tolerance to the preparation significantly in comparison to when the same preparation is administered without the nega¬ tively charged component. The improved tolerance may be reflected in the reduction or elimination of adverse systemic effects produced by a liposome preparation in which the electric charge component is absent. These systemic effects include, e.g., hemodynamic effects, such as hemodynamic depression, changes in blood pressure, heart rate, systolic pressure, diastolic pressure, or ECG intervals (PR, QRS, QT, QTc) , reflex tachycardia, prema¬ ture ventricular arrhythmias, increased respiration, sedation, and also changes in the blood cell and platelet counts, and death. It is recognized that this basic principle of improving tolerance to an otherwise deleterious structure by electrical-charge modification may be applicable to reagents other than liposomes. The present invention relates to an amount of electrical charge which is effective in, e.g., reducing hemodynamic effects, improving tolerance, or reducing adverse effects, of a liposome preparation. The effective amount of electrical charge may be added to the liposome preparation by forming liposomes in the presence of the effective amount of an electrically charged component or by adding the effective amount of charge to already formed liposomes. The undesired or adverse effects of a liposome preparation upon administration may be reduced or eliminated by the addition of an electrical charge, regardless of the composition of the administered liposomes. For example, a liposome preparation having systemic effects may already contain electrically-charged components. In this case, the introduction of an effective amount of an electrically-charged component may be in addition to the charged components which are already present in the liposome. The charged component introduced may be of the same type or charge already present in the liposome or of an altogether different type or charge. Thus, the present invention relates to the addition of a charge to a liposome already having charged components. The charged component may be incorporated into the liposome in various ways. By incorporating an electrically charged component into a liposome, the charged component may become, e.g. , a structural component of the lipid bilayer, and/or conjugated or non- covalently attached to a constituent of the liposome. For example, when liposomes are formed from a mixture comprising an effective amount of a negatively-charged stearic acid, the stearic acid may be incorporated into the lipid bilayer of the liposome. However, the effective amount of an electrically-charged component may also be added to an already formed liposome preparation, or an incompletely formed preparation, e.g., by conjugating or non-covalently attaching a charged component to the surface of the lipid bilayer. How the addition of the electrically-charged component to the liposome is accomplished is of minor interest. The decisive feature is that the electrically- charged component is present in the liposome in an amount which is effective, e.g., to improve host tolerance, reduce adverse effects, and/or reduce the hemodynamic effects produced by the administration of a liposome preparation. The electrical-charge can therefore be carried and incorporated into the liposome in a variety of ways, including, e.g., by the addition of charged components such as acids, e.g., stearic acid, and salts of these components such as ammonium salts; by coupling charged moieties to components of the lipid bilayer; and by the addition of bilayer-forming substances with charges, either coupled covalently or adhering by any other force. While the invention is not bound by any theory, typically a liposome according to the present invention presents an electrical-charge on the outer surface of its bilayer. For example, a liposome preparation comprising phosphatidyl choline is known to have an adverse hemodynamic effect. Its tolerance by the host organism may be improved by preparing a new batch of liposomes comprising an effective amount of an electrically-charged component in addition to the phosphatidyl choline, e.g., the effective amount of electrical charge may be carried by stearic acid. Alternatively, the charge may be introduced into liposomes which are already formed. For example, an electrical-charge may be introduced to the outer (exterior) surface of the lipid bilayer by chemical modification, or by covalently or noncovalently adding a charged moiety to its surface. The addition of a charge to a liposome preparation may also permit higher amounts of the preparation to be administered to the host organism without producing dele- terious effects normally attributed to liposome admini¬ stration. This is significant in that larger dosages of the agent encapsulated in the liposome may be attained. The addition of a charge to a liposome may also improve the stability and storage of the liposome preparation, e.g. , at temperatures greater than room temperature, such as 30° or 40°C. The nature of the effective amount of electrically charge component may be either positive or negative, as long as the desired effect is achieved, e.g., to improve tolerance, reduce adverse effects, and/or reduce the hemodynamic effects produced by administration of a liposome preparation. A negative charge is preferred. The charge may be, e.g., ionic or electronegative. In principle, once it is known that a particular liposome preparation is not tolerated by the host animal, e.g., as manifested by undesirable systemic affects, a charged component may be selected and then incorporated into the liposome preparation for the purpose of reducing or eliminating the side effects associated with its administration. By preparing a series of liposome preparations having various amounts and kinds of charge, it would be routine to administer and determine how much and what kind of charge is effective in, e.g., reducing hemodynamic effects and improving host tolerance. The amount of effective electrical charge may be carried by a molecule or a charge carrier. The molecule may have one or more charged moieties, e.g., a zwitterion. The charge may be located at any position on the charge carrier. The choice of the charge carrier will depend on various factors; e.g., availability, biocompatability, and desired effect. The charge carrier may be selected for other properties in addition to its ability to carry the desired charge. For example, the charge carrier may also function as a targeting agent, i.e., directing the liposome selectively to a desired tissue or cell type in the host organism. The charge carrier may be, e.g., biomolecule, a polymer, a lipid, a protein, a nucleic acid, a carbohydrate, a synthetic molecule, or any molecule or molecular structure which may be incorporated into the liposome bilayer and which possesses an effective amount of electrical charge. By incorporated the molecule into the liposome bilayer, the molecule may become structurally part of the liposome vesicle, e.g., integrated into the lipid bilayer, or peripherally attached to the liposome surface, or as a lipid constituent, itself. Additionally, small compounds with an additional acidic group can be coupled to an "anchor" such as cholesterol, phosphatidyl choline, fatty acid etc. sitting in the bilayer. The charge, preferably negative, can also be carried by amino acids or sugar acids coupled to any moiety able to adhere to the bilayer (cholesterol, phosphatidyl choline, fatty acid, etc.). The charge carrier may also be any lipid which carries a charge, e.g., including, sterols, fatty acids, glycerol esters, sphingosine, terpenes, or generally lipids, e.g., see TEXTBOOK OF CLINICAL CHEMISTRY (1986) , N.W. Tietz, editor, W.B. Saunders Co., Chapter 7, pp. 829-900. Stearic acid is a preferred example of a fatty acid having a negative electrical charge which may be employed in the present invention in an effective amount, e.g., to reduce hemodynamic effects of a parenterally administered liposome preparation. Other fatty acids may include, e.g., palmitic acid, arachidonic acid, linolic acid, linoleic acid, and/or mixtures thereof. The charge carrier may also be a protein which, e.g., is capable of integrating into the lipid bilayer or attaching peripherally to the surface. The charge of the protein may be produced by amino acids, carbohydrate linkages, or non-protein groups linked or joined to the protein. The protein may also be a protein having a lipid moiety. A protein may be chosen because of its desired charge and also because of its ability to target liposomes to a specific tissue. The amount of charge to be incorporated into the liposome may be that amount which is effective to reduce the undesired systemic effects such as hemodynamic depression and/or tolerance. The amount of charge may be varied by changing the molar ratio between the charged component and the other constituents of the liposome. For example, a charged component may be added to a liposome comprising phosphatidyl choline by the addition of stearic acid. The amount of charge in the liposome may be achieved by adjusting the molar ratio between the two constituents to a desired amount; e.g. , phosphatidyl choline/stearic acid, 9:1. The improved tolerance by the host to a liposome preparation and the consequent elimination or reduction in systemic affects may be produced by the incorporation of a charged component into the liposome vesicle. This improvement, when mediated by changes to the cell surface caused by the charged component, may also be produced by using agents or methods which modify surfaces. For example, the already formed liposomes having a deleterious systemic affect, may be treated enzymatically or chemically to alter the surface properties of the liposome and improve its tolerance when administered. Chemical modification may include e.g. , reacting the liposome reducing agents, oxidizing agents, or coupling charged components to its surface. These methods are accomplished conventionally but to achieve the reduction or elimination of adverse effects as revealed here. The liposomes may be prepared according to methods which are conventional in the art, e.g., as reviewed in W086/00238. Additionally, liposomes may be prepared according to EP69307; U.S. Pat. No. 5,110,475 which, e.g., describes a process for the production of an aqueous dispersion involving removal of liquid(s) from an optionally multiphase liquid mixture by means of membrane distillation; and W086/00238 which is described as an extrusion techniques for producing liposomes having substantially a unimodal and defined size distribution. When a liposome is prepared with an electrically- charged component, it is possible that the electrically- charged moiety may be oriented in the liposome in several different directions; e.g., facing the interior of the liposome vesicle and facing the exterior at the interface between aqueous medium and the lipid. Liposomes having the electrical charge on the exterior are preferred, e.g., the exterior surface. It may be desirable to separate, e.g., the "interior" charged from the "exterior" charged liposomes. This may be accomplished by any mean as known in the art, including column chromatography employing, e.g., charged or otherwise modified supports, electrophoresis, electrical separation processes, and other processes which separate structures based on charge, charge density, or charge orientation. Generally, the liposome preparations containing the electrically-charged component may be subjected to conditions which eliminate orientations which are not desirable for the purposes of the present invention. An active agent may be encapsulated in the liposomes according to conventional methodology. Thus, a liposome preparation having an effective amount of an electrically charged component may comprise or encapsulate an active agent. Encapsulation may be accomplished by methods which are known, e.g., by forming the liposome in the presence of an amount of an active agent under conditions in which the lipid bilayers form around and encapsulate the active agent, by introducing an active agent to an already formed liposome by fusion with a liposome, cell, or other structure comprising the active agent, e.g., such fusion may be carried out by PEG, electric current, lectins, and other well known methods in the art. In addition, liposomes having an active agent may be prepared according to WO/89593, e.g., by treating the liposomes in an aqueous medium under pressure conditions effective to reduce the order of the lipid arrangement therein to permit entry of the active agent and performing a pressure drop. An active agent may be encapsulated into a liposome preparation having an effective amount of an electrically charged component, including e.g. , conventional active agents such those used for therapy, including agents as listed in the Physicians Desk References, Medical Economics Data, 44th Edition, 1993 or Remington ' s Pharmaceutical Sciences , Mack Publishing Co., 18th Edition, 1990, antibodies, peptides, DNA, RNA, ribozymes, and oligonucleotides, prophylaxis, and diagnostics, e.g., contrast agents, such as iopromide, paramagnetic entities, ferromagnetic entities, radioisotopes, and other conventional structures or compounds. Such an active agent may be one that is used and recognized by the ordinary skilled worker in the field. The liposomes may be administered according to conventional methods. For example, a liposome preparation may be administered parenterally, e.g., intravenously by arterial or venous catheter, by injection with a syringe, however they may also be administered orally or through the intestines. A typical amount of liposome preparation is, e.g., 100-200 mg lipid/kg, this amount being dependent on how much of the agent is intended for administration and how much of it is encapsulated per liposome. The amount of liposome preparation which is to be administered may be determined routinely, according to methods which are known in the art. The liposomes may be administered in a single injection or it may be administered or infused over a period of time at a constant or variable rate, e.g., 2 ml/min for 30 minutes. Because the liposomes comprising the charged component are well tolerated by the host, infusion rates and amounts of liposome to be administered may be increased over the amounts that are typically used for liposome preparations which have adverse effects. The liposomes may be administered to any organism, e.g., mammals, such as humans, rats, mice, dogs, cats, rabbits, cows, horses, but also birds, amphibians, and reptiles. The purpose of administration may be to diagnose, treat, medicate, sedate, or prevent disorders but it may also be used in conjunction with the development of animal models to evaluate the safety and efficacy of drugs, contrast media, antibodies, and other agents for their eventual use in humans. The method of the present invention of e.g., reducing hemodynamic effects, improving host tolerance, or reducing adverse effects, associated with liposome administration, may be accomplished in any organism, such as those listed above. The liposomes can be used for any diagnostic or therapeutic use, including e.g., MRI, Ultrasound, and Nuclear Medicine. Without further elaboration, it is believed that one skilled in the art can, using the preceding description, utilize the present invention to its fullest extent. The following preferred specific embodiments are, therefore, to be construed as merely illustrative, and not limita¬ tive of the remainder of the disclosure in any way whatsoever. In the foregoing and in the following examples, all temperatures are set forth uncorrected in degrees Celsius and unless otherwise indicated, all parts and percentages are by weight. The entire disclosures of all applications, patents and publications, cited above and below, are hereby incorporated by reference. E X A M P L E S Example 1 The preparation process for iopromide liposomes by the "ethanol vaporation method" is depicted in Figure 1. An ethanolic solution of the membrane lipids (standard = 92.5 mg/ml phosphatidylcholine/cholesterol/stearic acid 4:5:1 molar) is mixed with an iopromide solution which is obtained by diluting aliquots of an Ultravist® 370 solu¬ tion with 20 mM tromethamine buffer, pH 7.4 to give a final iodine concentration of 46.25 mg/ml. After the mixing period, ethanol is removed from the mixture under reduced pressure until a white liposomal suspension is obtained. This suspension is then filtered (5 μm and 1.2 μm) , filled into vials and lyophilized to produce a stable form for storage. Prior to application, such lyophilizates are first rehydrated with 4 ml 135 mM mannitol solution per g of dry substance. The resulting liposomal suspension is then filtered (20 μm glass-fiber pre-filter and 5 μm CN main filter) to remove non-liposomal structures. These structures are sling- or loop-shaped bilayer formations in part larger than 10 μm. It has been shown that these structures are removed by the above filtration step. The suitability of this preparation process for re- producible production of larger amounts of liposomes has also been demonstrated. Preparation of 16 batches (1000 ml liposomal suspension each) resulted in encapsulation of 42.1 ± 4.9% and a mean diameter of 458 ± 55 nm in the resuspended material (2 ml 135 mM mannitol solution per g lyophilizate) . The process is potentially suited to performance under aseptic conditions. The solutions employed to produce the liposomes can be ultra- or sterile-filtered. The composition of a resuspended lyophilizate (4 ml 135 mM mannitol solution per g lyophilizate) is listed below (without ranges) : iopromide: phosphatidylcholine (PC) DL-α-tocopherole: cholesterol (CH) : stearic acid (SS) : tromethamine: 1 N HC1: disodium edetate: mannitol: water f.i. : A molar lipid mixture of PC/CH/SS 4:5:1 resulted in the highest degree of encapsulation (42.5 ± 3.7%, n = 6) as well as the smallest mean diameter (439 ± 87 nm) in the resuspended material. These values are in close agreement with the values that were obtained when the batch size was increased 10-fold (see 3.1). The molar ratio of three membrane components as well as the type and quality of each lipid component were optimized regarding these target parameters. The use of other non-ionic monomer contrast agents (iopamidol, iohexol and ZK 119095) rather than iopromide did not prove superior with regard to iodine encapsula¬ tion. When the non-ionic dimer iotrolan was encapsu¬ lated, higher weight ratios (mg iodine/mg lipid) were achieved. This compound was excluded from further studies, however, because of unacceptably long retention of liposomal iotrolan in the liver and spleen of test animals (biodistribution studies) and concomitant pathological changes in the liver. In the ethanol-evaporation process an iodine/lipid ratio of 1:1 in the original solutions (see Fig. 1) proved the most suitable as regards optimal use of the lipid (i.e., with the highest possible iodine to lipid weight ratio) in the final preparation. Of the resuspen- sion media tested, 135 mM mannitol resulted in the high¬ est encapsulation values in comparison with bidistilled water, various buffer solutions and mannitol solutions of lower concentrations. Example 2 For an initial stability study, a batch of liposome lyophilizate was produced under aseptic laboratory conditions. The resuspended liposomes were characterized regarding size (QUELS) , iodine encapsulation, pH and microscopic appearance. The microbial status was not determined. According to the results obtained in this study (six months' storage), liposomal lyophilizates (PC/CH/SS 4:5:1) can be stored at temperatures up to 30°C without perceivable quality changes. After 6 months at 40°C, however, a significant increase in size and a decrease in pH were observed in the resuspended material. These changes were accompanied by an unpleasant odor coming from the resuspended material. In a stability study of the (unfiltered) resuspended material, no significant changes were observed in size (QUELS) , encapsulation or pH over a period of 24 h. Possible changes in the microscopic appearance of the preparation were not monitored in this study, however; and so, the microscopic appearance of a resuspended lyophilizate was monitored separately both prior to and after filtration over a time-period of 8 hours. No significant changes were observed in either of the materials examined. Example 3 (A) Methods (i) Surgical Procedures Six to seven days prior to study, 6 normal healthy adult mongrel dogs of either sex were selected for similarity in weight (14.3-15.6 kg) and anesthetized with 35 mg/kg, iv pentobarbital sodium, shaved, and prepared for sterile surgery by washing the area of the intended incision twice with Betadine® surgical scrub, and then painting the area with Betadine® solution. The dogs were intubated using a cuffed endotracheal tube and mechanically respired with room air using a Harvard animal respirator (model 607) , placed in dorsal recumbency on the surgical table and draped for surgery using sterile techniques. A small ("1 inch) incision was made between the manubrium of the sternum and the greater tubercle of the right humerus. Using blunt dissection, the right omocervical artery and right jugular vein were isolated for cannulation. The artery was cannulated first using Tygon microbore tubing (0.05" ID, #S-54-HL) . The catheter was secured with 2-0 silk suture and exteriorized via a subdermal tunnel to the nape of the neck. In a similar fashion, the right jugular vein was catheterized using Tygon microbore tubing (0.04" ID, #S- 54-HL) and exteriorized to the nape of the neck also. Both arterial and venous catheters were filled with sterile, heparinized (600 U/mL) saline, sutured and painted with betadine to reduce infection. The exposed catheters were secured in a small plastic pouch and enclosed in a cotton-mesh collar reinforced with adhesive tape. Dogs were administered standard intramuscular injections of Combiotic to reduce the risk of infection and allowed to recover from surgery. (ii) Experimental Procedures On the day of study, the dogs were brought into the laboratory two at a time in individual mobile slings. The dogs were shaved at the ankles and instrumented with lead II electrocardiographic (ECG) leads using surface electrodes and conducting jelly. The specialized collars were removed, blood was withdrawn from each catheter and the catheters were flushed with sterile saline. The arterial catheter was attached to a pressure transducer and calibrated using a mercury manometer. A 10 mL sample of venous blood was collected from the venous catheter using sterile Sarstedt monovette tubes containing EDTA. Following an equilibration period of 20-40 minutes, dogs were administered an infusion of DMPC/DPPC, prepared by the interdigitation/fusion method (See WO 91/10422; EP 510096) , or DPPC at 2 mL/minute for "25 minutes. Parameters monitored were direct arterial blood pressure and lead II surface ECG. (iϋ) Drug Administration DMPC/DPPC or DPPC liposome vesicles were admini¬ stered intravenously as an opaque suspension at a rate of 2 mL/minute in a concentration of 1 mg/kg/ L until a total dose of 50 mg/kg was administered. All doses were loaded into sterile 60 mL syringes and infused using a Razel infusion pump. Any remaining suspension in the catheter was flushed into the dog using sterile saline at the same infusion rate. (iv) Statistical Analysis Selected CBC counts after vesicle administration were compared to basal values using a paired sample t-test. Differences were considered significant when p < 0.05. (B) Results The effects of DMPC/DPPC or DPPC infusions on heart rate (HR) , mean blood pressure (MAP) , systolic pressure (SP) , diastolic pressure (DP) , ECG intervals (PR, QRS, QT, QTc) , and selected blood cell count values are shown in tables 1-4. The complete blood cell count (CBC) values are contained in the appendix. (i) Blood Pressure Intravenous administration of DMPC/DPPC or DPPC vesicles caused a marked decrease in systolic, diastolic, and mean arterial pressure. (ii) Systolic Pressure (Table 1) Systolic pressure fell an average of 37% during infusion of DMPC/DPPC. An even greater fall in systolic pressure was observed during the infusion of DPPC vesicles with systolic pressure dropping 55%. While systolic pressures returned to near baseline levels in only one dog from each group during observation period, this pressure fall was determined to be transient as systolic pressures were within normal physiological levels within 30 minutes following vesicle administration. (iii) Diastolic Pressure (Table 2 ) Diastolic pressure fell 39% in the DMPC/DPPC group and 48% in the DPPC group during the vesicle infusion. One dog from the DMPC/DPPC group maintained control levels of diastolic pressure during the infusion and 1 dog from the DPPC group demonstrated minimal effects on diastolic pressure. On the average, the effect of liposome vesicles on diastolic pressure was transient, as mean diastolic pressures returned to near control levels within 30 minutes following vesicle administration. (iv) Mean Pressure (Table 3) The effect of liposome vesicles on mean arterial pressure was clearly more marked in the DPPC group with lower and more prolonged falls in pressure. The dogs in the DMPC/DPPC group experienced an average fall in mean pressure of 38% during infusion, but returned to near control levels by the end of the infusion. Dogs re¬ ceiving DPPC vesicles, however, recorded a 52% fall in mean pressure during infusion and did not recover to near control levels until 30 minutes following the end of the infusion. (v) Heart Rate (Table 4) The effect of liposome vesicles on heart rate in conscious dogs was variable. While mean heart rate values increased in both groups during administration of liposome vesicles, this reflects a pronounced tachycardia recorded from only one dog in each group. The other 2 dogs in each group maintained a heart rate equal to or lower than control during the infusion and the 45 minute observation period that followed. (vi) Electrocardiographic Intervals (Table 5) The infusion of DMPC/DPPC or DPPC liposome vesicles had no physiologically marked effect on ECG intervals. No changes were observed in PR or QRS intervals (Table 3A and 3B, pg. 13) . Minor increases in QT (Table 3C, pg. 14) and QTc (Table 3D, pg. 14) were recorded but did not reflect a major physiologic effect sufficient to be significant. (vii) CBC Values (Table 6) The infusion of DMPC/DPPC liposome vesicles increased white cell count significantly and lowered circulating platelets slightly, red cell count and hematocrit were not effected (Table 4A, pg. 15) . The infusion of DPPC vesicles also had little effect on red cell count and hematocrit, however did significantly lower platelet count while lowering white cell count slightly (Table 4B, pg. 15) . (C) Discussion The present study, in which conscious dogs were monitored for acute hemodynamic and ECG changes, showed the intravenous infusion of DMPC/DPPC and DPPC liposome vesicles caused a sudden and significant drop in blood pressure resulting in a variety of physiologic reactions including lethargy, and mild sedation in all dogs with emesis occurring in only one dog. No physiologically significant changes were noted in ECG parameters; however, white blood cell counts and circulating platelet counts were effected. The majority of these overt reactions could be attributed to the transient, but significant, decrease in blood pressure resulting from a possible anaphylactic reaction to the test substances. Accompanying such a reaction would be changes in white blood cell and circulating platelet counts. Example 4 (A) Methods (i) Surgical Procedures Six to seven days prior to study, 5 normal healthy adult mongrel dogs of either sex were selected for similarity in weight (13.7-14.1 kg) and anesthetized with 35 mg/kg, iv pentobarbital sodium, shaved, and prepared for sterile surgery by washing the area of the intended incision twice with Betadine® surgical scrub, then painting the area with Betadine® solution. The dogs were intubated using a cuffed endotracheal tube and mechanically respired with room air using a Harvard animal respirator (model 607) , placed in dorsal recumbency on the surgical table and draped for surgery using sterile techniques. A small (1 inch) incision was made between the manubrium of the sternum and the greater tubercle of the right humerus. Using blunt dissection, the right omocervical artery and right jugular vein were isolated for cannulation. The artery was cannulated first using Tygon microbore tubing (0.05" ID, /S-54-HL) . The catheter was secured with 2-0 silk suture and exteriorized via a subdermal tunnel to the nape of the neck. In a similar fashion, the right jugular vein was catheterized using Tygon microbore tubing (0.04 inch ID, #S-54-HL) and also exteriorized to the nape of the neck. Both arterial and venous catheters were filled with sterile, heparinized (600 U/mL) saline (1 mL) and capped with sterile caps. The 2 incisions were closed with silk suture and painted with Betadine® to reduce the risk of infection. The exposed catheters were secured in a small plastic pouch and enclosed in a cotton-mesh collar reinforced with adhesive tape. Dogs were administered standard intramuscular injections of Combiotic to further reduce the risk of infection and allowed to recover from surgery. (ϋ) Experimental Procedures On the day of study, the dogs were brought into the laboratory in individual mobile slings. The dogs were shaved at the ankles and instrumented with lead II electrocardiographic (ECG) leads using surface electrodes and conducting jelly. The specialized collars were re¬ moved, blood was withdrawn from each catheter and the catheters were flushed with sterile saline. The arterial catheter was attached to a pressure transducer and calibrated (0-200 mmHg) using a mercury manometer. A 10 mL sample of venous blood was collected from the venous catheter using sterile Sarstedt monovette tubes contain¬ ing EDTA. Following an equilibration period of 20-40 minutes, dogs were administered an infusion of EPC at 2 mL/minute for 25 minutes. Parameters monitored were direct arterial blood pressure and lead II surface ECG. (iii) Drug Administration EPC liposome vesicles were administered intraven¬ ously as an opaque suspension at a rate of 2 mL/minute in a concentration of 1 mg/kg/mL until a total dose of 50 mg/kg was administered. All doses were loaded into sterile 60 mL syringes and infused using a Razel infusion pump. Any remaining suspension in the catheter was flushed into the dog using sterile saline at the same infusion rate to assure that the total dose was administered to each dog. (B) Results The effects of EPC liposome vesicle infusions on heart rate (HR) , mean blood pressure (MAP) , systolic pressure (SP) , diastolic pressure (DP) , ECG intervals (PR, QRS, QT, QTc) , and complete blood count values (CBC) are described below. Peak changes in blood pressure and heart rate occurred during the infusion of liposome vesicles. (i) Blood Pressure Intravenous administration of EPC liposome vesicles caused a marked decrease in systolic, diastolic, and mean arterial pressure in 3 of the 5 dogs tested. (ii) systolic Pressure (Table 7) Systolic pressure fell an average of 36% in a group of 5 dogs. Those dogs experiencing acute hemodynamic depression (n = 3) had systolic pressure dogs ranging from 50% to 62% below basal levels. These changes in systolic pressure were transient as systolic pressure values returned to normal levels by the end of the EPC infusion. (iii) Diastolic Pressure (Table 8) Diastolic pressure fell 36% among the 5 dogs tested. Those 3 dogs which experienced a marked decrease in blood pressure had diastolic pressure drops ranging from 56% to 61% below basal levels. While these drops in pressure were severe, they were also transient as pressure values returned to near basal levels by the end of the infusion of the EPC vesicles. (iv) Mean Pressure (Table 9) The infusion of EPC liposome vesicles had no ob- served effect in 2 of the 5 dogs tested; however, 3 of the 5 dogs tested demonstrated marked falls in mean arterial pressure. The average fall in mean pressure was 35%, but those dogs with marked hemodynamic depression experienced mean pressure drops ranging from 57% to 61% below basal levels. While these drops in pressure were severe, they were also transient as pressure values returned to near basal levels by the end of the infusion of the EPC vesicles. (v) Heart Rate (Table 10) The effect of EPC liposome vesicles on heart rate in conscious dogs was closely related to physiologic re¬ sponse to falls in blood pressure. Those dogs which were compromised hemodynamically by the EPC infusion demon¬ strated a profound reflex tachycardia during EPC infusion followed by brief bradycardia, ith heart rate returning to basal levels in all dogs by 30 minutes post EPC infu¬ sion. The 2 dogs which did not experience hemodynamic compromise during EPC infusion maintained relatively constant heart rates throughout the study. (vi) Electrocardiographic Intervals (Table 11. A-D) The infusion of EPC liposome vesicles had no physio¬ logically significant effect on ECG intervals. No changes were observed in QRS duration. Changes occurring in PR, QT and QTc intervals were related to heart rate changes that occurred in the 3 dogs overtly affected by the EPC infusion. None of these changes were determined to be of physiologic significance due to their transient nature. In one dog, spontaneous arrhythmias (in the form of isolated premature ventricular contractions) occurred during the EPC infusion. Examples are shown in Figure 2. (vii) Mean CBC Values (Table 12) The infusion of EPC liposome vesicles did not effect red blood cell count or hematocrit. White blood cell counts were not consistent. Platelet counts were clearly depleted in those 3 dogs which experienced hemodynamic depression. One of the dogs not experiencing hemodynamic depression (USDA #97684) did experience a platelet depletion, but not below normal range values. (C) Discussion Intravenous administration of EPC liposome vesicles in conscious dogs resulted in hemodynamic depression. The platelet depletion observed in this study paralleled a similar effect noted in the previous study. In both studies, platelet depletion seemed closely related to severe falls in blood pressure resulting from liposome infusion. The overt reactions observed during liposome infusion (platelet depletion, marked hypotension, and reflex tachycardia) are most likely related to an anaphylactic response. The spontaneous arrhythmias observed in one dog receiving EPC vesicles may be a response to the sudden hypotensive effect with a reflex activation of catecholamines. Exam le 5 Liposomes according to the present invention, comprising the electrically-charged component stearic acid (preparations SA 504/02079 and SA 504/300591) , were administered to healthy dogs. (A) Methods (i) Surgical Procedures Six to seven days prior to study, normal healthy adult mongrel dogs of either sex were selected for simi- larity in weight (12.3-15.1 kg) and anesthetized with 35 mg/kg, iv pentobarbital sodium, shaved, and prepared for sterile surgery by washing the area of the intended inci¬ sion twice with Betadine® surgical scrub, then painting the area with endotracheal tube and mechanically respired with room air using a Harvard animal respirator (model 607) , placed in dorsal recumbency on the surgical table and draped for surgery using sterile techniques. A small (1 inch) incision was made between the manubrium of the sternum and the greater tubercle of the right humerus. Using blunt dissection, the right omocervical artery and right jugular vein were isolated for cannulation. The artery was cannulated first using Tygon microbore tubing (0.05" ID, #S-54-HL) . The catheter was secured with 2-0 silk suture and exteriorized via a subdermal tunnel to the nape of the neck. In a similar fashion, the right jugular vein was catheterized using Tygon microbore tubing (0.05" ID, /S-54-HL) and also exteriorized to the nape of the neck. Both arterial and venous catheters were filled with sterile, heparinized (600 U/mL) saline (1 mL) and capped with sterile painted with Betadine® to reduce infection. The exposed catheters were secured in a small plastic pouch and enclosed in a cotton-mesh col¬ lar reinforced with adhesive tape. Dogs were admini¬ stered standard intramuscular injections of Combiotic to reduce the risk of infection and allowed to recover from surgery. (ii) Experimental Procedures On the day of study, the dogs were brought into the laboratory and placed in individual mobile slings. The dogs were shaved at the ankles and instrumented with lead II electrocardiographic (ECG) leads using surface electrodes and conducting jelly. The specialized collars were removed, blood was withdrawn from each catheter and the catheters were flushed with sterile saline. The arterial catheter was attached to a pressure transducer and calibrated using a mercury manometer. A 10 mL sample of venous blood was collected from the venous catheter using sterile Sarstedt monovette tubes containing EDTA. Following an equilibration period of 20-30 minutes, dogs were administered an infusion of either SA 504/020791 or SA 504/300591 at approximately 2 mL/minute for exactly 30 minutes. Parameters monitored were arterial blood pressure and lead II surface ECG. Heart rate was determined from the ECG tracing. (iϋ) Drug Administration SA 504/020791 or SA 504/300591 iodine-containing liposome vesicles were administered intravenously as an opaque suspension at a rate of approximately 2 mL/minute at a final volume of 4.2 mL/kg and 4.5 mL/kg, respec- tively. The infusion rate was adjusted such that the total infusion time was 30 minutes. All doses were loaded into sterile syringes and infused using a Razel infusion pump. Any remaining suspension in the catheter was flushed into the dog using sterile saline at the same infusion rate. The test substances were prepared according to the following procedure: SA 504/020791: 17.6 mLs of mannitol solution (135 mM) was added to each vial of the test sub¬ stance and allowed to sit for at least 10 minutes. Vials were then shaken vigorously and allowed to sit for ano¬ ther 10 minutes. After repeating the shaking procedure, the suspensions were checked visually for agglomerates (if agglomerates were present, the above procedure of shaking the vials was repeated) . Suspensions were then drawn into 60 mL syringes and filtered through a sterile filter apparatus supplied by Schering AG. Each animal received 4.5 mL/kg. SA 504/300591: The procedure is exactly as outlined above except that 18.0 mLs mannitol solution (135 mMO were used per vial and each animal received 4.2 mL/kg. (iv) Statistical Analysis Hemodynamic parameters, ECG intervals, and selected complete blood counts were compared to pre-treatment basal values using a paired sample t-test. Differences were considered significant when p < 0.05. (v) Liposome Preparation SA 504/020791 and SA 5041/300591 were prepared according to the following instructions. A solution 1 was prepared by dissolving 54.6 gms of lipoid S100, 33.0 gms of cholesterol, and 4.9 gms of stearic acid in 9.5 ml of 96.5% ethanol. 2000 ml of a solution 2 was prepared by combining 250 ml of iopromide solution with 9.5 ml of 20 mM Tris HCl, pH 7..5, and 9.5 ml of 96% ethanol. While stirring, solution 1 was mixed with solution 2. The ethanol was removed under vacuum and the remaining solution was freeze-dried. 5 gm of freezed-dried material was resuspended in 8.8 ml of 135 mmol mannitol. The resuspended liposome preparation was typically injected at 300 mg iodine/kg. (B) Results The effects of SA 504/020791 or SA 504/300591 infu¬ sions on heart rate (HR) , mean blood pressure (MAP) , systolic pressure (SP) , diastolic pressure (DP) , ECG intervals (PR, QRS, QT, QTc) , and selected blood cell count values are shown in Tables 13-16. (i) Blood Pressure (Table 13) Intravenous administration of SA 504/020791 or SA 504/300591 vesicles produced no significant changes in systolic, diastolic or mean arterial pressures. As can be seen in Table 13C, control mean arterial pressure for dogs treated with SA 504/300591 was 104 ± 6 and remained relatively unchanged throughout the duration of the experiment. Similar results were obtained with the test substance SA 504/020791. (ii) Heart Rate The liposome vesicle preparations had no significant effect on heart rate in conscious dogs. Of the five dogs treated with SA 504/300591, three dogs had a decrease in heart rate during the infusion and two had an increase. Thus, no consistent pattern was seen. In one animal receiving SA 504/020791 (Dog #10003) , the control heart was particularly high (164 bpm) . This animal was highly reactive to any stimuli in the room (i.e., laboratory personnel) , but eventually seemed to quiet down as the experiment progressed. This was most likely due to acclimatization and not a drug-induced effect. (iii) Electrocardiographic Intervals The infusion of either test agent had no physiolo¬ gically marked effect on ECG intervals. No changes were observed in PR, QRS, QT, or QTC intervals. (vii) CBC Values The infusion of either test agent had no significant effect on RBC or WBC counts. Although the WBC values were not significantly affected, the trend appears that WBC will be increased following administration of the test agents. If one pools the data from both groups (i.e., n = 8), then statistical significance is achieved for elevating WBC (p = 0.026). In 5 experiments, platelet values before and after the test substances were categorized as either "adequate," "increased," or "decreased." In all 5, the platelet values were characterized as "adequate." In the remaining 3 experiments, values for platelet counts (thousands/mL) are as follows: Treatment Dog # Control 45' S*.504/300591 10174 435 376 S^ 504/020791 10003 198 243 SA_ 504/020791 98701 210 298 (C) Discussion Previous studies on liposome preparations conducted in conscious dogs with DMPC/DPPC, DPPC, and EPC liposome vesicles demonstrated a dramatic, transient hypotensive response in "40-60% of the animals tested. The present study, in which conscious dogs were monitored for acute hemodynamic and ECG changes, showed the intravenous infusion of SA 504/300591 and SA 504/020791 liposome preparations produced no significant changes in hemodynamic variables (blood pressure and heart rate) , ECG intervals, or RBC and WBC values, and are therefore well tolerated and free of adverse hemodynamic effects. Example 6 Two liposome formulations were prepared and tested in a dog model for hemodynamic effects. Preparation l: Liposomes which only contained phos¬ phatidyl choline at a concentration of ca. 25 mg/ml. Preparation 2: Liposomes with phosphatidyl choline and stearic acid. The molar content was ca. 9:1 at a total lipid concentration of ca. 70 mg/hl. These preparations were infused intravenously into five dogs each at a dose level of ca. 50 mg lipid per kg body weight (preparation 1) and ca. 30 mg lipid per kg (preparation 2) . The following effects were observed in the dogs. Preparation l: Transient fall in blood pressure associated with a significant reflex tachycardia in 3 of the 5 animals tested. Other observed effects in these 3 animals included: premature ventricular arrhythmias dur¬ ing the infusion and briefly following the administration in 1 dog and increased respiration in the form of panting in the 2 other dogs. There were no physiologically sig¬ nificant changes in the ECG intervals in any of the dogs. Platelet counts fell in 4 of 5 dogs receiving the prepa¬ ration. Those dogs experiencing hemodynamic depression seemed somewhat sedated and had the most consistent platelet depletion when compared with those dogs who did not experience decreases in blood pressure. Preparation 2: Intravenous administration of this formulation had no adverse hemodynamic effects in any of the five animals treated. Example 7 In this example, the cardio-hae odynamic effects of liposomes prepared from a saturated uncharged phospho- lipid DSPC alone or in a combination with a negatively charged phosolipid DSPG (9:1) were examined. Preparation of liposomes: The liposomes which were used in this study were prepared by a continuous high pressure extrusion method. Briefly, an ethanolic solution of the respective lipid or lipid mixtures was deposited on the wall of a round bottom flask. The resulting lipid film was dispersed in 300 mM mannitol solution to give a lipid concentration of approximately 100 mg/ml. The resulting MLV dispersions were subsequently extruded using a high pressure extrusion apparatus (Maximator® model HPE 10.0 - 250, Schmidt, Kranz & Co., Zorge, Germany). Each batch was sequentially extruded 10 times over two stacked polycarbonate membranes (Nucleopore, Tiibingen, Germany) of decreasing pore sizes (1.0 - 0.4 and 0.1 μm) to give a total of 30 filter passages for each preparation. At the end of the extrusion process the obtained liposomal suspensions were filtered through sterile filter holders (0.4 or 0.2 μm pore size cellulose acetate, Sartorius, Gδttingen, Germany) into sterile glass vials which were stoppered under aseptic conditions. Lipid Substances All phospholipids were stored below - 20°C and used without further purification. Distearoylphosphatidylcholine (DSPC- batches LP-04-013- 114219 and -116298) and Distearoylphosphatidyl-glycerol (DSPG - batch LP 04-017-115751) were obtained from Sygena, Liestal, Switzerland. Liposome size was determined by photon correlation spectroscopy (PCS) using a submicron particle-sizer, autodilute™ model 370, Nicomp Instr. Corp., Santa Barbara, CA, USA. Osmolality [mOsm/kg] of samples and pH values were determined with an automatic freezing point osmometer (Knauer, Berlin, Germany) and a pH meter 761 Calimatic (Knick, Berlin, Germany) , respectively. Animal experiments Eighteen male rats (strain: Han-Wistar, breeder: Schering SPF, standard feeding and housing conditions) , with a body mass of 360 to 430 g were randomly sorted into three groups with n=6 animals per group. The rats were anaesthetized with pentobarbital, 60 mg/kg intraperitoneally. The trachea was cannulated to facilitate spontaneous respiration. Body temperature was maintained at 38 + 0.5 C by means of a heated operating table and a heating lamp. Left ventricular pressure and blood pressure in the femoral artery were recorded via polyethylene filled catheters connected to Statham pressure transducers. Cardiac output was determined by the thermodilution method (thermistor catheter in the abdominal aorta, injection of ice cold saline into the right atrium) . The animals were heparinized with an i.v. bolus of 400 U/kg. The rats were set up to allow recording/calculation of the following parameters: BPsyst. (systolic blood pressure, mmHg) , BPmean (mean blood pressure, mmHg) , BPdiast. (diastolic blood pressure, mmHg) , HR (heart rate, 1/min) , CO (cardiac output/100 g body mass, ml/min/100 g) , LVEDP (left ventricular enddiastolic pressure, mmHg) , TPR (total peripheral resistance, dyne*s*cm' 5 10 3 ) , ECG (electrocardiogram, ms) and dP/dt max (maximum rate of left ventricular pressure rise, mmHg/s) . Surgery and instrumentation were followed by a 30- min equilibration period, at the end of which the pre- treatment values were determined. Data were recorded for 45 min after administration of the liposomes or mannitol (volume control) . Each animal received one treatment only. Rats were infused with a liposome dose containing a total amount of 300 mg lipid/kg body weight and at an injection rate of 100 mg lipid/kg/min into the left femoral vein. Controls received an identical volume of 300 mM mannitol solution. The investigated liposome formulations were made from DSPC or DSPC/DSPG. Statistics Mean percentage changes of the haemodynamic data versus prevalues were calculated (except for LVEDP, which was expressed as absolute change from pre-drug baseline values) . Data were compared with the control group. The Bartlett-test was carried out to test for equal variances and one way analysis of variance (ANOVA) was used to demonstrate statistical differences. Calculations of statistical significance were performed using Student's t-test. A p value below 0.05 was considered statis¬ tically significant. Values in the figures are expressed as mean percentage changes from prevalue + SEM. Results Control experiments Mannitol 300 mM (volume control) did not cause cardio-haemodynamic parameters to significantly differ from prevalues. BPmean increased maximally by 1.8% (Fig. 2), BPsyst. by 2.3% and PBdiast. decreased by -2.2% within the first 10 min post application. HR decreased by -3.4%. TPR (Fig. 3) and LVEDP (Fig. 4) responded with a short time increase by 5.5% and 2.8 mmHg, respectively (Tab. 17). Contractility decreased by -3.3%. DSPC liposomes The use of the saturated DSPC resulted in neutral, gel-state liposomes which led to marked cardiac and haemodynamic disturbances when infused at a dose of 300 mg lipid/kg into rats. BPmean and TRP were significantly decreased by -53.7% and -46.3%, respectively (Figs. 2 and 3). Contractility was reduced by -34.7% 5 min after the start of infusion. During this time LVEDP (Fig. 4) was significantly increased (p<0.05). All these effects were accompanied by severe cardiac arrhythmia. There were ventricular extrasystolies, ectopic beats, AV conduction delays and failure of conduction to the ventricle. However, HR and CO were only slightly affected during the first 10 min post application. At the end of this 10 min period BP and contractility returned to baseline values. HR, however, started to increase slightly above baseline (about 10%) and contractility too. Until the end of the observation period at 45 min p.a., contractility was increased to +66.3% above baseline (p<0.01). TPR, however, did not return to baseline by the end of the observation time. DSPC/DSPG liposomes The addition of a negatively charged phospholipid (10 mol%) to DSPC reduced the severity of the cardio- haemodynamic side effects of DSPC significantly. BP and TPR decreased after a short transient increase to -33.3% and -35.5% of prevalue, respectively (Figs. 2 and 3). Contractility declined by -14.3% during the first 10 min post application. It returned to normal values and was increased at 30 min p.a. HR also responded with an increase at later time points. Electrocardiographic changes occurred in four of six rats during the liposome infusion. The effects were interpreted as conducted and nonconducted atrial premature beats in three animals and in one animal as complex arrhythmia produced by paired atrial premature beats. All ECG-effects disappeared after the end of the infusion. Thus, the addition of a negative charge to DSPC liposomes significantly improved the cardio-hemodynamic side effects of the preparation in rats. The preceding examples can be repeated with similar success by substituting the generically or specifically described reactants and/or operating conditions of this invention for those used in the preceding examples. From the foregoing description, one skilled in the art can easily ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions. TABLE 1 A. Systolic Pressure υ_ TABLE 2 B. Diastolic Pressure 00 c Mean Pressure TAllLE o 'IΛBLL 5 PR Interval B. QRS Duration K) TΛBLI- 5 (Cont'd) c. QT Interval D. QT Interval ■ -^ TAB LI DMPC/DPPC -P- B. DPPC TΛBLΓ. 7 SYSTOLIC PRESSURE (mmllg) IN EPC-TREΛTED DOGS TΛ I.L 8 DIASTOLIC PRESSURE (mmHg) VALUES IN EPC VESICLE TREATED DOGS I ' ΛI. LE 9 MEAN ARTERIAL PRESSURE (mmHg) VALUES IN EPC TREATED DOGS TABLE 10 HEART RA TE VALUES (Beats/Minute) IN EPC VESICLE TREATED DOGS 1-1 σ '.- ' PR (msec) VALUES IN EPC TREATED DOGS B. QRS (msec) VALUES IN EPC TREATED DOGS (Conc ' d . QT (msec) VALUES IN EPC TREATED DOGS QT C (msec) VALUES IN EPC TREATED DOGS l r.D i BLOOD COUNTS IN EPC TRE.ATED DOGS TABLE 13 Λ Systolic Prc33ure Uι B. Diastolic Pressure Ui C. Mean Pressure l TΛI.I.I: !-'• -J * Animal sleeping during iπfu3ion ** Personnel entering room TΛI-l.l. 15 Λ. PR Intorval ι 00 D. QRS Duration z> C. QT Interval Oι O D. QTc Interval σ TΛI.U_ 16 A. RBC (million/mL) o B a en • σ c tύ n Table 17 O ^. n«β, • p<0.u5," p<0-01 l t " p<0.001 l Student* t-tβat Maximum percentage change of haemodynamic and cardiac parameierβ during the flrβt 10 min after start of infusion of liposomes made from different Iipldt (300 mg lipld/kg, 100 mg lipld/kg/mln) into the the femoral vein.

Claims: WHAT IS CLAIMED IS; 1. A method of reducing the hemodynamic effects of a parenterally administratable liposome preparation which, if administered to a mammal would cause hemodynamic effects, comprising: incorporating in the liposome preparation prior to administration an amount of an electrically charged component effective to reduce said hemodynamic effects, wherein said amount is in addition to any amount of said component in said liposome preparation. 2. A method of improving a host's tolerance for a parenterally administratable liposome preparation comprising: incorporating in the liposome preparation an amount of an electrically charged component, in addition to any amount of such component in said liposome preparation, effective to improve said tolerance. 3. In a method of administering an active agent encapsulated in a liposome preparation having adverse systemic effects, the improvement comprising incorporating in the liposome preparation prior to administration an amount of an electrically charged component effective in reducing said adverse systemic effects, wherein said amount is in addition to any amount of said component in said liposome. 4. In a method of imaging a tissue of a host organism comprising administering a contrast media encapsulated in a liposome preparation having adverse systemic effects, the improvement comprising incorporating in the liposome preparation prior to administration an amount of an electrically charged component effective in reducing said systemic effects, wherein said amount is in addition to any amount of said component in said liposome. 5. A method of reducing the hemodynamic effects of a parenterally administratable liposome preparation comprising: administering a liposome preparation comprising an amount of an electrically charged component effective in reducing hemodynamic effects associated with liposome administration. 6. The method of claim 1, wherein the electrically charged component has a negative charge. 7. The method of claim 1, wherein the electrically charged component is a fatty acid. 8. The method of claim 1, wherein the electrically charged component is stearic acid. 9. The method of claim 1, wherein the liposome further comprises an active agent. 10. The method of claim 1, wherein the liposome further comprises a contrast agent. 11. The method of claim 1, wherein the contrast agent is iopromide. 12. In a liposome which does not contain an electrically charged component and which has adverse systemic effects when administered to a mammal, the improvement comprising incorporating in said liposome an amount of an electrically charged component effective in reducing said systemic effects, wherein said amount is in addition to any amount of said component in said liposome. 13. The liposome of claim 12, wherein the electrically charged component has a negative charge. 14. The liposome of claim 12, wherein the electrically charged component is a fatty acid. 15. The liposome of claim 12, wherein the electrically charged component is stearic acid. 16. The liposome of claim 12, further comprising an active agent. 17. The liposome of claim 12, further comprising a contrast agent. 18. In a liposome which comprises an electrically charged component and which has adverse systemic effects when administered to a mammal, the improvement comprising incorporating in said liposome an amount of an electrically charged component effective in reducing said systemic effects, wherein said amount is in addition to any amount of said component in said liposome. 19. The liposome of claim 18, wherein the electrically charged component has a negative charge. 20. The liposome of claim 18, wherein the electrically charged component is a fatty acid. 21. The liposome of claim 18, wherein the electrically charged component is stearic acid. 22. The liposome of claim 18, further comprising an active agent. 23. The liposome of claim 18, further comprising a contrast agent. 24. In a method of administering a therapeutic agent encapsulated in a liposome preparation having adverse systemic effects, the improvement comprising incorporating in the liposome preparation prior to administration an amount of an electrically charged component effective in reducing said adverse systemic effects, wherein said amount is in addition to any amount of said component in said liposome. 25. A method according to claim 1, wherein the electrically charged component incorporated into said liposome preparation is stearic acid and it is incorporated in an amount such that the liposome comprises phosphatidylcholine, cholesterol, and stearic acid in a molar ratio of about 4:5:1, respectively. 26. A method according to claim 1, wherein the electrically charged component incorporated into said liposome preparation is stearic acid and it is incorporated in an amount such that the liposome comprises phosphatidylcholine and stearic acid in a molar ratio of about 9:1, respectively. 27. A method according to claim 1, wherein the electrically charged component incorporated into said liposome preparation is DSPG and it is incorporated in an amount such that the liposome comprises DSPC and DSPG in a molar ratio of about 9:1, respectively. 28. The method of claim 1, wherein the electrically charged component is DSPG. 29. The method of claim 5, wherein the electrically charged component is DSPG. 30. The method of claim 12, wherein the electrically charged component is DSPG. 31. The method of claim 24, wherein the electrically charged component is DSPG. 32. A method of reducing the hemodynamic effects of a parenterally adininistrable liposome preparation which, if administered to a mammal, would cause hemodynamic effects, comprising: identifying a liposome preparation having adverse hemodynamic effects, and incorporating into said liposome preparation an amount of an electrically charged component effective to reduce said hemodynamic effects, said amount being in addition to any amount of said component in said liposome preparation.

Inventors: KRAUSE WERNER, SACHSE ANDREAS, SULLIVAN MARK, KRAUSE, WERNER, SACHSE, ANDREAS, SULLIVAN, MARK
Applicants: KRAUSE WERNER, SACHSE ANDREAS, SCHERING AG, SULLIVAN MARK, KRAUSE, WERNER, SACHSE, ANDREAS, SCHERING AKTIENGESELLSCHAFT, SULLIVAN, MARK
Application year: 1994
Main classification: A61K9
IPC codes: A61K9, A61K49